Comparative analysis of osteoderms across the lizard body.

Osteoderms (ODs) are mineralized tissue embedded within the skin and are particularly common in reptiles. They are generally thought to form a protective layer between the soft tissues of the animal and potential external threats, although other functions have been proposed. The aim of this study was to characterize OD variation across the lizard body. Adults of three lizard species were chosen for this study. After whole body CT scanning of each lizard, single ODs were extracted from 10 different anatomical regions, CT scanned, and characterized using sectioning and nanoindentation. Morphological analysis and material characterization revealed considerable diversity in OD structure across the species investigated. The scincid Tiliqua gigas was the only studied species in which ODs had a similar external morphology across the head and body. Greater osteoderm diversity was found in the gerrhosaurid Broadleysaurus major and the scincid Tribolonotus novaeguineae. Dense capping tissue, like that reported for Heloderma, was found in only one of the three species examined, B. major. Osteoderm structure can be surprisingly complex and variable, both among related taxa, and across the body of individual animals. This raises many questions about OD function but also about the genetic and developmental factors controlling OD shape.

Effect of atropine and propofol on the minimum anaesthetic concentration of isoflurane in the freshwater turtle Trachemys scripta (yellow-bellied slider).

OBJECTIVE: To determine if the administration of atropine would reduce the measured minimum anaesthetic concentration of isoflurane (MACisoflurane) in freshwater turtles - the yellow-bellied slider (Trachemys scripta scripta). STUDY DESIGN: Paired, blinded, randomized, prospective studies of 1) the effect of atropine in isoflurane anaesthetized freshwater turtles (T. scripta scripta) and 2) the effect of atropine in yellow-bellied sliders in which anaesthesia was induced with propofol and maintained with isoflurane. ANIMALS: T. scripta scripta (n = 8), female, adult. METHODS: Atropine (2 mg kg-1) or an isovolumetric control injection of saline was administered intraperitoneally 15 minutes prior to induction of anaesthesia with isoflurane. Individual MACisoflurane was then determined by end-tidal gas analysis in a bracketing design by an experimenter blinded to the administered drug, with a 2 week washout period. The experiment was repeated, with atropine (2 mg kg-1) or saline administered intravascularly in combination with propofol for anaesthetic induction. Linear mixed modelling was used to determine the effects of atropine and propofol on the individual MACisoflurane. Data are presented as mean ± standard deviation. RESULTS: Premedication with atropine significantly reduced MACisoflurane (p = 0.0039). In isoflurane-induced T. scripta scripta, MACisoflurane decreased from 4.2 ± 0.4% to 3.3 ± 0.8% when atropine had been administered. Propofol as an induction agent had a MAC-sparing effect (p < 0.001) such that MACisoflurane following propofol and a control injection of saline was 2.3 ± 1.0%, which decreased further to 1.5 ± 0.8% when propofol was combined with atropine. CONCLUSIONS AND CLINICAL RELEVANCE: Atropine, presumably by inhibiting parasympathetically mediated pulmonary artery constriction, decreases right-to-left cardiac shunting and the MACisoflurane in yellow-bellied sliders, and thereby may facilitate control of inhalant anaesthesia. Propofol can be used for induction of anaesthesia and reduces the required concentration of inhaled anaesthesia assessed 1.5 hours following induction.

The role of cranial osteoderms on the mechanics of the skull in scincid lizards.

Osteoderms (ODs) are calcified organs formed directly within the skin of most major extant tetrapod lineages. Lizards possibly show the greatest diversity in ODs morphology and distribution. ODs are commonly hypothesized to function as a defensive armor. Here we tested the hypothesis that cranial osteoderms also contribute to the mechanics of the skull during biting. A series of in vivo experiments were carried out on three specimens of Tiliqua gigas. Animals were induced to bite a force plate while a single cranial OD was strain gauged. A finite element (FE) model of a related species, Tiliqua scincoides, was developed and used to estimate the level of strain across the same OD as instrumented in the in vivo experiments. FE results were compared to the in vivo data and the FE model was modified to test two hypothetical scenarios in which all ODs were (i) removed from, and (ii) fused to, the skull. In vivo data demonstrated that the ODs were carrying load during biting. The hypothetical FE models showed that when cranial ODs were fused to the skull, the overall strain across the skull arising from biting was reduced. Removing the ODs showed an opposite effect. In summary, our findings suggest that cranial ODs contribute to the mechanics of the skull, even when they are loosely attached.

Obesity prolongs induction times in reptiles.

Obesity is common in captive reptiles, and reptiles are increasingly popular as companion animals and in physiological research. Obesity may present a challenge during surgical procedures using inhalation anaesthesia, as the long induction time due to the low reptilian metabolism may increase anaesthetic accumulation in the adipose tissues. This study investigated the impact of obesity on induction and recovery times from inhaled anaesthesia. The temporal change in the partial pressure of isoflurane in different tissues was predicted using a multi-compartment model. Furthermore, as right-to-left shunting can delay anaesthetic uptake and washout, we included an assessment of the combination of cardiac shunting and obesity. The model predictions indicate a clear increase in time to reach 90% equilibration of administered anaesthetic in the brain (T90) of obese non-shunting (lean 47 min, obese >100 min) and shunting (lean 81 min, obese >100 min) reptiles. The combination of obesity and shunting doubled the time to acquisition of mean anaesthetic concentration (a measure used to plan anaesthesia) from 8 min to 19 min. Adipose blood flow highly affected whether the body type had an impact on induction time, with low adipose blood flow abolishing the effect of body type. As T90 was never reached within 100 min with both the obese reptiles, it was not possible to conclude on the effect of obesity on recovery times within this study. Care should therefore be taken when anaesthetising obese reptiles for surgical purposes, to ensure adequate anaesthetic depth is attained, and recovery monitored closely.

Unravelling the structural variation of lizard osteoderms.

Vertebrate skin is a remarkable organ that supports and protects the body. It consists of two layers, the epidermis and the underlying dermis. In some tetrapods, the dermis includes mineralised organs known as osteoderms (OD). Lizards, with over 7,000 species, show the greatest diversity in OD morphology and distribution, yet we barely understand what drives this diversity. This multiscale analysis of five species of lizards, whose lineages diverged ∼100-150 million years ago, compared the micro- and macrostructure, material properties, and bending rigidity of their ODs, and examined the underlying bones of the skull roof and jaw (including teeth when possible). Unsurprisingly, OD shape, taken alone, impacts bending rigidity, with the ODs of Corucia zebrata being most flexible and those of Timon lepidus being most rigid. Macroscopic variation is also reflected in microstructural diversity, with differences in tissue composition and arrangement. However, the properties of the core bony tissues, in both ODs and cranial bones, were found to be similar across taxa, although the hard, capping tissue on the ODs of Heloderma and Pseudopus had material properties similar to those of tooth enamel. The results offer evidence on the functional adaptations of cranial ODs, but questions remain regarding the factors driving their diversity. STATEMENT OF SIGNIFICANCE: Understanding nature has always been a significant source of inspiration for various areas of the physical and biological sciences. Here we unravelled a novel biomineralization, i.e. calcified tissue, OD, forming within the skin of lizards which show significant diversity across the group. A range of techniques were used to provide an insight into these exceptionally diverse natural structures, in an integrated, whole system fashion. Our results offer some suggestions into the functional and biomechanical adaptations of OD and their hierarchical structure. This knowledge can provide a potential source of inspiration for biomimetic and bioinspired designs, applicable to the manufacturing of light-weight, damage-tolerant and multifunctional materials for areas such as tissue engineering.

The influence of assisted ventilation and recumbency on cardiorespiratory physiology in the anesthetized freshwater turtle Trachemys scripta scripta.

The use of assisted ventilation is required in anesthetized reptiles as their respiratory drive is lost at surgical depths of anesthesia. The minute volume of the assisted ventilation influences arterial blood gases and acid-base regulation. Meanwhile, the ventilatory pattern may also affect hemodynamics in chelonians, which, given their large capacity for cardiac shunts, may impact the efficacy of the ventilation in terms of gas exchange. Hence, there is a need for primary information on the influence of assisted ventilation on chelonian physiology, and we, therefore, performed a randomized study into the effects of recumbency and maximum airway pressure on pressure-cycled ventilation in nine female Trachemys scripta scripta. Pronounced effects of ventilation pressure on arterial PCO2 and pH regardless of recumbency were revealed, whilst dorsal recumbency led to a larger Arterial-alveolar (A-a) O2 difference, suggesting compromised pulmonary gas exchange. Plasma [Na+] and [K+] balance was also significantly correlated with maximum airway pressure. Computed tomography (CT) scanning at a range of end-inspiratory pressures and ventral and dorsal recumbencies in eight T. scripta scripta showed that lung volumes increase with maximum ventilatory pressure, while recumbency did not influence volume at pressures above 5 cmH2O. Static compliance of the lungs was influenced by recumbency at neutral pressures. In conclusion, dorsal recumbency reduces pulmonary efficacy during positive pressure ventilation and tends to lower lung volume when ventilation is not provided. However, lung volumes and function - even in dorsal recumbency - can be adequately supported by assisted ventilation, and an end inspiratory pressure of 10 cmH2O at 4 breaths min-1 provided the most physiologically appropriate ventilation of anesthetized T. scripta scripta.

Ectothermy and cardiac shunts profoundly slow the equilibration of inhaled anaesthetics in a multi-compartment model.

The use of inhalational anaesthesia is ubiquitous in terrestrial vertebrates. Given the dependence of these agents on delivery by the cardiorespiratory system, we developed a new computational model predicting equilibration of inhaled anaesthetics in mammalian and ectotherm conditions including the ability of reptiles to maintain vascular shunts. A multi-compartment model was constructed from simultaneously-solved equations, verified by comparison to the literature for endo and ectotherm physiology. The time to 90% equilibration of anaesthetic in arterial blood (t90) is predicted and used to compare anaesthetics and physiologies. The five to tenfold lower cardiac output and minute ventilation of ectothermic vertebrates is predicted to slow equilibration times by five to ten times leading to 90% equilibration in ectotherm arterial blood of over 200 min, compounded by reduction in body temperature, and the extent of right-to-left vascular shunts. The impact of these findings is also influenced by the solubility coefficient of the anaesthetic, such that at net right-to-left shunt fractions of over 0.8, sevoflurane loses the advantage of faster equilibration, in comparison with isoflurane. We explore clinical strategies to regulate anaesthetic uptake in ectotherms by managing convectional flow especially by supportive ventilation and reduction of the right-to-left shunt.

Extracting physiological information in experimental biology via Eulerian video magnification.

BACKGROUND: Videographic material of animals can contain inapparent signals, such as color changes or motion that hold information about physiological functions, such as heart and respiration rate, pulse wave velocity, and vocalization. Eulerian video magnification allows the enhancement of such signals to enable their detection. The purpose of this study is to demonstrate how signals relevant to experimental physiology can be extracted from non-contact videographic material of animals. RESULTS: We applied Eulerian video magnification to detect physiological signals in a range of experimental models and in captive and free ranging wildlife. Neotenic Mexican axolotls were studied to demonstrate the extraction of heart rate signal of non-embryonic animals from dedicated videographic material. Heart rate could be acquired both in single and multiple animal setups of leucistic and normally colored animals under different physiological conditions (resting, exercised, or anesthetized) using a wide range of video qualities. Pulse wave velocity could also be measured in the low blood pressure system of the axolotl as well as in the high-pressure system of the human being. Heart rate extraction was also possible from videos of conscious, unconstrained zebrafish and from non-dedicated videographic material of sand lizard and giraffe. This technique also allowed for heart rate detection in embryonic chickens in ovo through the eggshell and in embryonic mice in utero and could be used as a gating signal to acquire two-phase volumetric micro-CT data of the beating embryonic chicken heart. Additionally, Eulerian video magnification was used to demonstrate how vocalization-induced vibrations can be detected in infrasound-producing Asian elephants. CONCLUSIONS: Eulerian video magnification provides a technique to extract inapparent temporal signals from videographic material of animals. This can be applied in experimental and comparative physiology where contact-based recordings (e.g., heart rate) cannot be acquired.

Magnetic resonance imaging (MRI) reveals high cardiac ejection fractions in red-footed tortoises (Chelonoidis carbonarius).

The ejection fraction of the trabeculated cardiac ventricle of reptiles has not previously been measured. Here, we used the gold standard clinical methodology - electrocardiogram-gated flow magnetic resonance imaging (MRI) - to validate stroke volume measurements and end diastolic ventricular blood volume. This produced an estimate of ejection fraction in our study species, the red footed tortoise Chelonoidis carbonarius (n=5), under isoflurane anaesthesia of 88±11%. After reduction of the prevailing right-to-left intraventricular shunt through the action of atropine, the ejection fraction was 96±6%. This methodology opens new avenues for studying the complex hearts of ectotherms, and validating hypotheses on the function of a more highly trabeculated heart than that of endotherms, which have lower ejection fractions.

The effects of endogenous and exogenous catecholamines on hypoxic cardiac performance in red-bellied piranhas.

Catecholamines protect the fish heart during hypoxia. However, the humoral adrenergic stress response may only be invoked in extremis. We investigated the hypothesis that endogenous (e.g., neuronal) myocardial catecholamines may also impact cardiac performance during hypoxia in a hypoxia-tolerant tropical fish, the red-bellied piranha (Pygocentrus nattereri). First, we measured endogenous tissue catecholamines and in vitro catecholamine release from piranha myocardium using ultraperformance liquid chromatography. Ventricle homogenates contained detectable levels of both adrenaline (7.27 ng/g) and noradrenaline (14.48 ng/g), but only noradrenaline was released from ventricular tissue incubated in Ringer's solution. Noradrenaline released in this assay was not affected by hypoxia but was promoted by the catecholamine releasing agent tyramine. Our second series of experiments explored cardiac contractile performance in vitro using tyramine, exogenous noradrenaline or adrenaline, and propranolol (a ß-adrenoceptor antagonist). In ventricular strip preparations, ß-adrenergic blockade with propranolol had no effects on twitch force or contraction kinetics in either normoxia or hypoxia, confirming that spontaneous endogenous catecholamine release did not impact cardiac performance. However, in the absence of propranolol, tyramine mimicked the positive inotropic effect of noradrenaline (10 µM) during hypoxia, although adrenaline was capable of generating larger effects. Our results suggest that, although it is not spontaneously released, inducible endogenous noradrenaline release may have a significant ß-adrenoceptor-dependent impact on hypoxic performance in the fish heart.

CARDIOVASCULAR EFFECTS OF ALFAXALONE AND PROPOFOL IN THE BULLFROG, LITHOBATES CATESBEIANUS.

Alfaxalone is becoming a popular anesthetic for nonmammalian vertebrates, but the physiological effects of its administration remain largely unknown in these taxa. Therefore, the cardiovascular responses to a clinically relevant dose of alfaxalone (10 mg/kg) are reported in the bullfrog ( Lithobates catesbeianus), following intramuscular (IM) and intravascular (IV) administration (via a femoral artery catheter) and compared with an IV dose of propofol, another parenteral GABA (γ-aminobutyric acid) agonist in common veterinary use as an induction agent. Heart rate (HR) and mean arterial blood pressure (MAP) (assessed by direct measurement from the catheter) are reported from under undisturbed conditions to assess both the direct effects of the drugs and the interaction with the stress of handling associated with IM injection of alfaxalone where IM administration is possible. Alfaxalone caused HR to increase significantly for over 45 min in both groups from a baseline of approximately 30 beats/min. This was significantly different from the lack of significant HR response on the IV administration of propofol. MAP increased in the peri-injection period with both routes of administration for alfaxalone but after IV use decreased significantly from 10 min following administration. Propofol did not affect blood pressure after 5 min from injection. Assessment of immobilization following intramuscular injection of alfaxalone in a pilot study was in accordance with the literature, as it provided no antinociception as a sole agent but did produce sedation and loss of righting reflex.

Venous pressures and cardiac filling in turtles during apnoea and intermittent ventilation.

The amount of blood pumped by the heart (cardiac output) must be matched to the amount of blood returning to the heart (venous return), but the factors determining cardiac filling are sparsely understood in ectothermic vertebrates. Stroke volume is affected by heart rate along with central and peripheral venous pressures. In the present study, we investigated the heart rate dependency of cardiac filling in turtles, along with the changes in venous pressures that accompany ventilation. Experimental reductions in heart rate of anaesthetised turtles (Trachemys scripta) by the specific bradycardic agent zatebradine (2-3 mg kg-1) resulted in an elevation of stroke volume that compensated cardiac output. By contrast, in spontaneously ventilating turtles, stroke volume remained constant, even during the transitions from the pronounced bradycardia during breath-hold diving to the accelerated heart rate associated with spontaneous ventilation. Ventilation was associated with pronounced decreases in visceral, pericardial and central venous pressure, all of which became sub-ambient (especially during inspiration) and may provide a powerful 'suctional' element to cardiac filling. In addition, mean circulatory filling pressure, an index of vascular capacitance and the peripheral driving pressure for venous return, was increased by infusion of adrenaline (2.5 µg kg-1). Together these data demonstrate that changes in both central and peripheral venous pressures are key determinants of venous return that, in concert with direct regulation of the heart, contribute to the large scope for cardiac output in turtles.

A critical test of Drosophila anaesthetics: Isoflurane and sevoflurane are benign alternatives to cold and CO2.

Anaesthesia is often a necessary step when studying insects like the model organism Drosophila melanogaster. Most studies of Drosophila and other insects that require anaesthesia use either cold exposure or carbon dioxide exposure to induce a narcotic state. These anaesthetic methods are known to disrupt physiology and behavior with increasing exposure, and thus ample recovery time is required prior to experimentation. Here, we examine whether two halogenated ethers commonly used in vertebrate anaesthesia, isoflurane and sevoflurane, may serve as alternative means of insect anaesthesia. Using D. melanogaster, we generated dose-response curves to identify exposure times for each anaesthetic (cold, CO2, isoflurane and sevoflurane) that allow for five-minutes of experimental manipulation of the animals after the anaesthetic was removed (i.e. 5min recovery doses). We then compared the effects of this practical dose on high temperature, low temperature, starvation, and desiccation tolerance, as well as locomotor activity and fecundity of female flies following recovery from anaesthesia. Cold, CO2 and isoflurane each had significant or near significant effects on the traits measured, but the specific effects of each anaesthetic differed, and effects on stress tolerance generally did not persist if the flies were given 48h to recover from anaesthesia. Sevoflurane had no measureable effect on any of the traits examined. Care must be taken when choosing an anaesthetic in Drosophila research, as the impacts of specific anaesthetics on stress tolerance, behavior and reproduction can widely differ. Sevoflurane may be a practical alternative to cold and CO2 anaesthesia in insects - particularly if flies are to be used for experiments shortly after anesthesia.

EVALUATION OF FEEDING BEHAVIOR AS AN INDICATOR OF PAIN IN SNAKES.

The necessity to prevent and manage pain in reptiles is becoming increasingly important, as their use in scientific research and popularity as exotic pets continues to rise. It was hypothesized that feeding behavior would provide an adequate indicator of pain perception in the ball python (Python regius). Normal feeding was defined the previous week, where a dead rodent was struck within 12 sec (n = 10). Eighteen pythons were randomly assigned to one of three treatments: anesthesia only (AO), chemical noxious stimulus (CS; capsaicin injection), or surgical noxious stimulus (SS; surgical incision). The time to strike was recorded 4 hr after the procedure and weekly during the subsequent 3 wk. Delayed feeding was observed in animals in the CS and SS groups, and normal feeding resumed after 1 and 3 wk, respectively. Spontaneous feeding remained uninterrupted for the AO group. These findings demonstrate feeding behavior as a potential model to assess pain in snakes.

When local anesthesia becomes universal: Pronounced systemic effects of subcutaneous lidocaine in bullfrogs (Lithobates catesbeianus).

Sodium channel blockers are commonly injected local anesthetics but are also routinely used for general immersion anesthesia in fish and amphibians. Here we report the effects of subcutaneous injection of lidocaine (5 or 50mgkg-1) in the hind limb of bullfrogs (Lithobates catesbeianus) on reflexes, gular respiration and heart rate (handled group, n=10) or blood pressure and heart rate via an arterial catheter (catheterized group n=6). 5mgkg-1 lidocaine did not cause loss of reflexes or change in heart rate in the handled group, but was associated with a reduction in gular respiratory rate (from 99±7 to 81±17breathsmin-1). 50mgkg-1 lidocaine caused a further reduction in respiratory rate to 59±15breathsmin-1, and led to a progressive loss of righting reflex (10/10 loss by 40min), palpebral reflex (9/10 loss at 70min), and contralateral toe pinch withdrawal (9/10 loss at 70min). Reflexes were regained over 4h. Systemic sedative effects were not coupled to local anti-nociception, as a forceps pinch test at the site of injection provoked movement at the height of the systemic effect (tested at 81±4min). Amphibians are routinely subject to general anesthesia via exposure to sodium channel blockers such as MS222 or benzocaine, however caution should be exercised when using local injectable lidocaine in amphibians, as it appears to dose-dependently cause sedation, without necessarily preventing local nociception for the duration of systemic effects.

Long-term surgical anaesthesia with isoflurane in human habituated Nile Crocodiles.

A suitable long-term anaesthetic technique was required for implantation of physiological sensors and telemetric devices in sub-adult Nile crocodiles (Crocodylus niloticus) to allow the collection of physiological data. Five Nile crocodiles with a median body mass of 24 kg were used. After manual capture, they were blindfolded and 0.2 mL (1 mg/mL) medetomidine was administered intramuscularly in four of the animals which had an estimated body mass between 20 kg and 30 kg. One crocodile with an estimated body mass of 50 kg received 0.5 mL. For induction, 5 mL propofol (10 mg/mL) was injected intravenously into the occipital sinus. Additional doses were given when required to ensure adequate anaesthesia. Anaesthesia was maintained with 1.5% isoflurane. Ventilation was controlled. Local anaesthesia was administered for surgical incision and external placement of the radio transmitter. Medetomidine was antagonised with atipamezole at the end of surgery. Median heart rate during surgery was 22 beats/min, at extubation 32 beats per min and 30 beats per min the following day at the same body temperature as under anaesthesia. Median body temperature of the animals increased from 27.3 °C to 27.9 °C during anaesthesia, as room temperature increased from 24.5 °C to 29.0 °C during surgery. Anaesthesia was successfully induced with intramuscular medetomidine and intravenous propofol and was maintained with isoflurane for the placement of telemetric implants. Intraoperative analgesia was supplemented with lidocaine infiltration. Perioperative physiological parameters remained stable and within acceptable clinical limits. Multiple factors appear to influence these variables during the recovery period, including residual anaesthetic effects, environmental temperature and physical activity.

The influence of mechanical ventilation on physiological parameters in ball pythons (Python regius).

Mechanical ventilation is widely recommended for reptiles during anesthesia, and while it is well-known that their low ectothermic metabolism requires much lower ventilation than in mammals, very little is known about the influence of ventilation protocol on the recovery from anesthesia. Here, 15 ball pythons (Python regius) were induced and maintained with isoflurane for 60min at one of three ventilation protocols (30, 125, or 250mlmin-1kg-1 body mass) while an arterial catheter was inserted, and ventilation was then continued on 100% oxygen at the specified rate until voluntary extubation. Mean arterial blood pressure and heart rate (HR) were measured, and arterial blood samples collected at 60, 80, 180min and 12 and 24h after intubation. In all three groups, there was evidence of a metabolic acidosis, and snakes maintained at 30mlmin-1kg-1 experienced an additional respiratory acidosis, while the two other ventilation protocols resulted in normal or low arterial PCO2. In general, normal acid-base status was restored within 12h in all three protocols. HR increased by 143±64% during anesthesia with high mechanical ventilation (250mlmin-1kg-1) in comparison with recovered values. Recovery times after mechanical ventilation at 30, 125, or 250mlmin-1kg-1 were 289±70, 126±16, and 68±7min, respectively. Mild overventilation may result in a faster recovery, and the associated lowering of arterial PCO2 normalised arterial pH in the face of metabolic acidosis.

Tachycardia in response to remote capsaicin injection as a model for nociception in the ball python (Python regius).

OBJECTIVES: To quantify the effect of subcutaneous (SC) capsaicin injection on heart rate (HR) in ball pythons (Python regius) and to assess the efficacy of two opioids (morphine and butorphanol) in modifying this response. STUDY DESIGN: Prospective, randomized, unmatched study. ANIMALS: Eleven mixed-sex, captive-bred ball pythons. METHODS: Snakes were randomly assigned to three groups (n = 6) by intramuscular premedication: 1) control: saline (0.9 mL); 2) morphine (10 mg kg(-1) ); and 3) butorphanol (10 mg kg(-1) ). Three snakes were tested twice and another two were tested three times in different treatments administered 1 month apart. Under isoflurane anaesthesia, snakes were instrumented with SC electrocardiogram (ECG) electrodes and an SC catheter for remote stimulus delivery. After recovery from anaesthesia, all snakes, in visual and audial isolation from the experimenter, received a sham stimulus of saline (0.4 mL) via the SC catheter. A nociceptive stimulus of SC capsaicin (3 mg in 0.2 mL saline with 7% Tween 80) was then applied by catheter at 7 hours after premedication. In a subset (n = 3), two sham injections (saline 0.2 mL) preceded the capsaicin treatment. HR was recorded via ECG, and changes in HR (ΔHR) from baseline were calculated for all stimulations. RESULTS: Capsaicin injection was associated with a significant increase in HR [peak ΔHR: saline group: 8.8 ± 7.1 beats minute(-1) ; capsaicin group: 21.1 ± 5.8 beats minute(-1) (p = 0.0055)] and integrated ΔHR as a function of time. The administration of morphine or butorphanol 7 hours prior to nociception failed to significantly reduce the peak and integrated ΔHR. Butorphanol caused marked, long-lasting sedation as assessed by muscle tone. CONCLUSIONS AND CLINICAL RELEVANCE: The HR response to an SC capsaicin injection can serve as a nociceptive model in P. regius. Morphine and butorphanol administration did not reduce HR response to capsaicin stimulation but produced significantly different effects on pre-stimulation HR and sedation.